A Preliminary Study On The Function Of Bovine ISG15 In BIV Lantent Infection

Lantent infection is the strategy how lentiviruses escape from immune sruveillance and immune clearance of the host. Innate immunity and adaptive immunity play important roles in defending the virus infection, while the interferon regulation is an important part of innate immunity. Interferon can stimulate the expression of hundrands of interferon stimulated genes (isgs). The over expression of interferon stimulated protein can inhibit the replication of viruses. The inhibition can lend lentivirus to establish latent infection. Latent infection of lentiviruses can be broken by many factors, including virus super-infection. Some virus super-infection with lentivirus can stimulate the replication of the lentivirus.In this work, we used BIV as a lentivirus model, and did a preliminary study on the function of bovine ISG15 (bISG15) in latent infection of BIV.ISG15 is a small molecular protein whose expression can be up-regulated by type I interferon. It also belongs to the family of ubiquitin-like modifiers. Like ubiquitin, ISG15 can be conjugated to other proteins to modify them. The process of ISG15 modification is called ISGylation. ISG15 and ISGylation play important roles in many processes including innate immunity, cancer and pregnancy. The amino acid sequences analysis shows that the cross-species conservation of ISG15 is not high. Recently, most studies on bovine ISG15 (bISG15) were focused on its role in pregnancy. However, studies about the functions that bISG15 is involeved in innate immune response or antiviral effect was rarely done.In order to relieve the relation between bISG15 and viruses which infected cattle, we established the RT-PCR assay system to detect the mRNA level of bISG15, the Western-blot assay to detect the expression level of bISG15 and reporting system of bISG15 gene promoter in vitro. These assay systems were used to detect the expression of bISG15 in FBLs which were used in the culture of BIV. Then, we did some research work in the relationship between bISG15 and BIV, and the infuence of BVDV or BHV-1 infection on bISG15 expression. The main findings and significance of this study are: 1. RT-PCR and Westen-blot assayes suggested that the basal expression of bISG15 in FBLs was too low to be detected, and the expression of bISG15 was induced by poly I:C or LPS. 2. Immunofluorescence assay and ChIP assay demonstrated that IRF-3 played a role in inducing the expression of bISG15 in FBLs. The Luciferase assay showed that the over-expression of bovine IRF-3 could active the promoter of bISG15 gene. 3. Infection of BIV can up-regulated the expression of bISG15 weakly in FBLs. 4. The replication of BIV in FBLs can partly repressed by bISG15. bISG15 plays a role in the establishment of latent infection of BIV. Meanwhile, bTat, which is the regulatory protein of BIV, seems to be involed in the repression. 5. BVDV or BHV-1 infection can repress the expression of bISG15. bICP0, which is the protein of BHV-1, can repress the activation of bISG15 gene promoter by IRF-3. BVDV or BHV-1 can help BIV replicating though repressing the interferon signal pathway.